scholarly journals Cobalamin (vitamin B12) biosynthesis: identification and characterization of a Bacillus megaterium cobI operon

1998 ◽  
Vol 335 (1) ◽  
pp. 159-166 ◽  
Author(s):  
Evelyne RAUX ◽  
Anne LANOIS ◽  
Martin J. WARREN ◽  
Alain RAMBACH ◽  
Claude THERMES
Keyword(s):  
Bacillus Megaterium ◽  
Genomic Library ◽  
Early Stage ◽  
Open Reading Frames ◽  
Biosynthetic Genes ◽  
Pseudomonas Denitrificans ◽  
Dna Fragment ◽  
Identification And Characterization ◽  
Synechocystis Sp

A 16 kb DNA fragment has been isolated from a Bacillus megaterium genomic library and fully sequenced. The fragment contains 15 open reading frames, 14 of which are thought to constitute a B. megaterium cobalamin biosynthetic (cob) operon. Within the operon, 11 genes display similarity to previously identified Salmonella typhimurium cobalamin biosynthetic genes (cbiH60, -J, -C, -D, -ET, -L, -F, -G, -A, cysGA and btuR), whereas three do not (cbiW, -X and -Y). The genes of the B. megaterium cob operon were compared with the cobalamin biosynthetic genes of Pseudomonas denitrificans, Methanococcus jannaschii and Synechocystis sp. Taking into account the presence of cbiD and cbiG, the absence of a cobF, cobG and cobN, -S and -T, it was concluded that B. megaterium, M. jannaschii and Synechocystis sp., like S. typhimurium, synthesize cobalamin by an anaerobic pathway, in which cobalt is added at an early stage and molecular oxygen is not required.

A transitional stage in the commitment of mesoderm to hematopoiesis requiring the transcription factor SCL/tal-1

Development ◽  
2000 ◽  
Vol 127 (11) ◽  
pp. 2447-2459 ◽  
Author(s):  
S.M. Robertson ◽  
M. Kennedy ◽  
J.M. Shannon ◽  
G. Keller
Keyword(s):  
Transcription Factor ◽  
Embryoid Body ◽  
Early Stage ◽  
Es Cells ◽  
Blast Cell ◽  
Stage Of Development ◽  
Erythroid Precursors ◽  
Identification And Characterization ◽  
Definitive Hematopoiesis

In this report, we describe the identification and characterization of an early embryoid body-derived colony, termed the transitional colony, which contains cell populations undergoing the commitment of mesoderm to the hematopoietic and endothelial lineages. Analysis of individual transitional colonies indicated that they express Brachyury as well as flk-1, SCL/tal-1, GATA-1, (beta)H1 and (beta)major reflecting the combination of mesodermal, hematopoietic and endothelial populations. This pattern differs from that found in the previously described hemangioblast-derived blast cell colonies in that they typically lacked Brachyury expression, consistent with their post-mesodermal stage of development (Kennedy, M., Firpo, M., Choi, K., Wall, C., Robertson, S., Kabrun, N. and Keller, G. (1997) Nature 386, 488–493). Replating studies demonstrated that transitional colonies contain low numbers of primitive erythroid precursors as well as a subset of precursors associated with early stage definitive hematopoiesis. Blast cell colonies contain higher numbers and a broader spectrum of definitive precursors than found in the transitional colonies. ES cells homozygous null for the SCL/tal-1 gene, a transcription factor known to be essential for development of the primitive and definitive hematopoietic systems, were not able to form blast colonies but did form transitional colonies. Together these findings suggest that the transitional colony represents a stage of development earlier than the blast cell colony and one that uniquely defines the requirement for a functional SCL/tal-1 gene for the progression to hematopoietic commitment.

scholarly journals Identification and Characterization of Four Autophagy-Related Genes That Are Expressed in Response to Hypoxia in the Brain of the Oriental River Prawn (Macrobrachium nipponense)

2019 ◽  
Vol 20 (8) ◽  
pp. 1856 ◽  
Author(s):  
Shengming Sun ◽  
Ying Wu ◽  
Hongtuo Fu ◽  
Xianping Ge ◽  
Hongzheng You ◽  
...  
Keyword(s):  
Developmental Stages ◽  
Open Reading Frames ◽  
Dependent Manner ◽  
Macrobrachium Nipponense ◽  
Oriental River Prawn ◽  
Adverse Environmental Conditions ◽  
The Brain ◽  
Identification And Characterization

Autophagy is a cytoprotective mechanism triggered in response to adverse environmental conditions. Herein, we investigated the autophagy process in the oriental river prawn (Macrobrachium nipponense) following hypoxia. Full-length cDNAs encoding autophagy-related genes (ATGs) ATG3, ATG4B, ATG5, and ATG9A were cloned, and transcription following hypoxia was explored in different tissues and developmental stages. The ATG3, ATG4B, ATG5, and ATG9A cDNAs include open reading frames encoding proteins of 319, 264, 268, and 828 amino acids, respectively. The four M. nipponense proteins clustered separately from vertebrate homologs in phylogenetic analysis. All four mRNAs were expressed in various tissues, with highest levels in brain and hepatopancreas. Hypoxia up-regulated all four mRNAs in a time-dependent manner. Thus, these genes may contribute to autophagy-based responses against hypoxia in M. nipponense. Biochemical analysis revealed that hypoxia stimulated anaerobic metabolism in the brain tissue. Furthermore, in situ hybridization experiments revealed that ATG4B was mainly expressed in the secretory and astrocyte cells of the brain. Silencing of ATG4B down-regulated ATG8 and decreased cell viability in juvenile prawn brains following hypoxia. Thus, autophagy is an adaptive response protecting against hypoxia in M. nipponense and possibly other crustaceans. Recombinant MnATG4B could interact with recombinant MnATG8, but the GST protein could not bind to MnATG8. These findings provide us with a better understanding of the fundamental mechanisms of autophagy in prawns.

scholarly journals Functional-Genomics-Based Identification and Characterization of Open Reading Frames Encoding α-Glucoside-Processing Enzymes in the Hyperthermophilic Archaeon Pyrococcus furiosus

2007 ◽  
Vol 74 (4) ◽  
pp. 1281-1283 ◽  
Author(s):  
Donald A. Comfort ◽  
Chung-Jung Chou ◽  
Shannon B. Conners ◽  
Amy L. VanFossen ◽  
Robert M. Kelly
Keyword(s):  
Glycoside Hydrolase ◽  
Bioinformatics Analysis ◽  
Pyrococcus Furiosus ◽  
Transcriptional Response ◽  
Open Reading Frames ◽  
Processing Enzymes ◽  
Response Information ◽  
Identification And Characterization ◽  
Reading Frames

ABSTRACT Bioinformatics analysis and transcriptional response information for Pyrococcus furiosus grown on α-glucans led to the identification of a novel isomaltase (PF0132) representing a new glycoside hydrolase (GH) family, a novel GH57 β-amylase (PF0870), and an extracellular starch-binding protein (1,141 amino acids; PF1109-PF1110), in addition to several other putative α-glucan-processing enzymes.

scholarly journals Identification and Characterization of Aspergillus nidulans Mutants Impaired in Asexual Development under Phosphate Stress

Cells ◽  
2019 ◽  
Vol 8 (12) ◽  
pp. 1520 ◽  
Author(s):  
Ainara Otamendi ◽  
Eduardo A. Espeso ◽  
Oier Etxebeste
Keyword(s):  
Transcription Factor ◽  
Aspergillus Nidulans ◽  
Genomic Library ◽  
Phenotypic Trait ◽  
Mutant Form ◽  
Multicopy Suppressor ◽  
Putative Transcription Factor ◽  
Recessive Mutant ◽  
Identification And Characterization

The transcription factor BrlA plays a central role in the production of asexual spores (conidia) in the fungus Aspergillus nidulans. BrlA levels are controlled by signal transducers known collectively as UDAs. Furthermore, it governs the expression of CDP regulators, which control most of the morphological transitions leading to the production of conidia. In response to the emergence of fungal cells in the air, the main stimulus triggering conidiation, UDA mutants such as the flbB deletant fail to induce brlA expression. Nevertheless, ΔflbB colonies conidiate profusely when they are cultured on a medium containing high H2PO4− concentrations, suggesting that the need for FlbB activity is bypassed. We used this phenotypic trait and an UV-mutagenesis procedure to isolate ΔflbB mutants unable to conidiate under these stress conditions. Transformation of mutant FLIP166 with a wild-type genomic library led to the identification of the putative transcription factor SocA as a multicopy suppressor of the FLIP (Fluffy, aconidial, In Phosphate) phenotype. Deregulation of socA altered both growth and developmental patterns. Sequencing of the FLIP166 genome enabled the identification and characterization of PmtCP282L as the recessive mutant form responsible for the FLIP phenotype. Overall, results validate this strategy for identifying genes/mutations related to the control of conidiation.

scholarly journals Identification and Characterization of a Novel Robigovirus Species from Sweet Cherry in Turkey

Pathogens ◽  
2019 ◽  
Vol 8 (2) ◽  
pp. 57 ◽  
Author(s):  
Kadriye Çağlayan ◽  
Vahid Roumi ◽  
Mona Gazel ◽  
Eminur Elçi ◽  
Mehtap Acioğlu ◽  
...  
Keyword(s):  
High Throughput Sequencing ◽  
Sweet Cherry ◽  
Prunus Avium ◽  
Open Reading Frames ◽  
Cherry Tree ◽  
Coding Regions ◽  
Sweet Cherries ◽  
Identification And Characterization ◽  
Reading Frames

High throughput sequencing of total RNA isolated from symptomatic leaves of a sweet cherry tree (Prunus avium cv. 0900 Ziraat) from Turkey identified a new member of the genus Robigovirus designated cherry virus Turkey (CVTR). The presence of the virus was confirmed by electron microscopy and overlapping RT-PCR for sequencing its whole-genome. The virus has a ssRNA genome of 8464 nucleotides which encodes five open reading frames (ORFs) and comprises two non-coding regions, 5′ UTR and 3′ UTR of 97 and 296 nt, respectively. Compared to the five most closely related robigoviruses, RdRp, TGB1, TGB2, TGB3 and CP share amino acid identities ranging from 43–53%, 44–60%, 39–43%, 38–44% and 45–50%, respectively. Unlike the four cherry robigoviruses, CVTR lacks ORFs 2a and 5a. Its genome organization is therefore more similar to African oil palm ringspot virus (AOPRV). Using specific primers, the presence of CVTR was confirmed in 15 sweet cherries and two sour cherries out of 156 tested samples collected from three regions in Turkey. Among them, five samples were showing slight chlorotic symptoms on the leaves. It seems that CVTR infects cherry trees with or without eliciting obvious symptoms, but these data should be confirmed by bioassays in woody and possible herbaceous hosts in future studies.

scholarly journals Postgenomic Scan of Metallo-β-Lactamase Homologues in Rhizobacteria: Identification and Characterization of BJP-1, a Subclass B3 Ortholog from Bradyrhizobium japonicum

2006 ◽  
Vol 50 (6) ◽  
pp. 1973-1981 ◽  
Author(s):  
Magdalena Stoczko ◽  
Jean-Marie Frère ◽  
Gian Maria Rossolini ◽  
Jean-Denis Docquier
Keyword(s):  
Bradyrhizobium Japonicum ◽  
Catalytic Efficiency ◽  
Open Reading Frames ◽  
Human Pathogens ◽  
Microbial Genomes ◽  
E Coli ◽  
Genes Encoding ◽  
And Function ◽  
Identification And Characterization

ABSTRACT The diffusion of metallo-β-lactamases (MBLs) among clinically important human pathogens represents a therapeutic issue of increasing importance. However, the origin of these resistance determinants is largely unknown, although an important number of proteins belonging to the MBL superfamily have been identified in microbial genomes. In this work, we analyzed the distribution and function of genes encoding MBL-like proteins in the class Rhizobiales. Among 12 released complete genomes of members of the class Rhizobiales, a total of 57 open reading frames (ORFs) were found to have the MBL conserved motif and identity scores with MBLs ranging from 8 to 40%. On the basis of the best identity scores with known MBLs, four ORFs were cloned into Escherichia coli for heterologous expression. Among their products, one (blr6230) encoded by the Bradyrhizobium japonicum USDA110 genome, named BJP-1, hydrolyzed β-lactams when expressed in E. coli. BJP-1 enzyme is most closely related to the CAU-1 enzyme from Caulobacter vibrioides (40% amino acid sequence identity), a member of subclass B3 MBLs. A kinetic analysis revealed that BJP-1 efficiently hydrolyzed most β-lactam substrates, except aztreonam, ticarcillin, and temocillin, with the highest catalytic efficiency measured with meropenem. Compared to other MBLs, BJP-1 was less sensitive to inactivation by chelating agents.

scholarly journals Characterization of ssfR andssgA, Two Genes Involved in Sporulation ofStreptomyces griseus

2000 ◽  
Vol 182 (19) ◽  
pp. 5521-5529 ◽  
Author(s):  
Hao Jiang ◽  
Kathleen E. Kendrick
Keyword(s):  
Stop Codon ◽  
Streptomyces Griseus ◽  
Large Family ◽  
Resistant Mutant ◽  
Open Reading Frames ◽  
Dna Fragment ◽  
Morphological Differences ◽  
White Mutant ◽  
Reading Frames

ABSTRACT In the presence of cefoxitin, which inhibits septum formation during sporulation, Streptomyces griseus is unable to sporulate, retaining the sonication sensitivity of nonsporulating hyphae. Cefoxitin- and sonication-resistant mutant SKK2600 was isolated and showed many morphological differences from its parental strain. A 3.6-kb DNA fragment that complemented the mutations of SKK2600 contained two open reading frames (ORFs), either of which could complement SKK2600. One ORF, designated ssfR, encoded a protein containing a potential DNA-binding helix-turn-helix motif close to its N terminus. SsfR is similar to members of a large family of transcriptional regulators, particularly IclR of Escherichia coli. The second ORF was identified as ssgA, a previously described sporulation gene from S. griseus (S. Kawamoto and J. C. Ensign, Actinomycetology 9:136–151, 1995). A point mutation of C to T seven nucleotides upstream of the UGA stop codon of ssfR was responsible for the phenotype of isolated mutant strain SKK2600. Surprisingly, this mutation should not change the primary structure of SsfR. The ssfR andssgA disruption mutants were constructed and showed the “white” mutant phenotype, with some growth medium dependence. In addition, the ssfR null mutant sporulated ectopically in phosphate starvation medium.

scholarly journals Cloning and Characterization of a New Site-Specific Methyl-Directed ElmI Endonuclease Recognizing and Cleaving C5-methylated DNA Sequence 5’-G(5mC)^NG(5mC)-3’

Acta Naturae ◽  
2016 ◽  
Vol 8 (1) ◽  
pp. 117-125 ◽  
Author(s):  
V. A. Chernukhin ◽  
D. A. Gonchar ◽  
M. A. Abdurashitov ◽  
O. A. Belichenko ◽  
V. S. Dedkov ◽  
...  
Keyword(s):  
Dna Sequence ◽  
Amino Acid Sequences ◽  
Open Reading Frames ◽  
Site Specific ◽  
Pcr Screening ◽  
Methylated Dna ◽  
Chromatographic Techniques ◽  
Dna Fragment ◽  
Reading Frames

Putative open reading frames of MD-endonucleases have been identified in Enterobacteria genomes as a result of the search for amino acid sequences homologous to MD-endonuclease BisI. A highly conserved DNA primary structure of these open reading frames in different genera of Enterobacteria (Escherichia, Klebsiella and Cronobacter) has allowed researchers to create primers for PCR screening, which was carried out on Enterobacteria DNA collected from natural sources. The DNA fragment, about 440 bp in length, was amplified by use of the genomic DNA of a wild E.coli LM N17 strain as a template and was inserted into the pMTL22 vector. Endonuclease activity was detected in an E.coli ER 2267 strain transformed with the obtained construction. A new enzyme named ElmI was purified by chromatographic techniques from the recombinant strain biomass. It was discovered that similarly to BisI this enzyme specifically cleaves the methylated DNA sequence 5-GCNGC- 3 before the central nucleotide N if this sequence contains two 5-methylcytosines. However, unlike BisI, ElmI more efficiently cleaves this sequence if more than two cytosine residues are methylated.

scholarly journals Identification and Characterization of a Gene Cluster for Synthesis of the Polyketide Antibiotic 2,4-Diacetylphloroglucinol from Pseudomonas fluorescens Q2-87

1999 ◽  
Vol 181 (10) ◽  
pp. 3155-3163 ◽  
Author(s):  
M. Gita Bangera ◽  
Linda S. Thomashow
Keyword(s):  
Genomic Dna ◽  
Plant Pathogens ◽  
Polyketide Synthase ◽  
Open Reading Frames ◽  
Fungal Plant Pathogens ◽  
Repressor Proteins ◽  
Flanking Regions ◽  
Identification And Characterization ◽  
Reading Frames

The polyketide metabolite 2,4-diacetylphloroglucinol (2,4-DAPG) is produced by many strains of fluorescent Pseudomonas spp. with biocontrol activity against soilborne fungal plant pathogens. Genes required for 2,4-DAPG synthesis by P. fluorescensQ2-87 are encoded by a 6.5-kb fragment of genomic DNA that can transfer production of 2,4-DAPG to 2,4-DAPG-nonproducing recipientPseudomonas strains. In this study the nucleotide sequence was determined for the 6.5-kb fragment and flanking regions of genomic DNA from strain Q2-87. Six open reading frames were identified, four of which (phlACBD) comprise an operon that includes a set of three genes (phlACB) conserved between eubacteria and archaebacteria and a gene (phlD) encoding a polyketide synthase with homology to chalcone and stilbene synthases from plants. The biosynthetic operon is flanked on either side by phlEand phlF, which code respectively for putative efflux and regulatory (repressor) proteins. Expression in Escherichia coli of phlA, phlC, phlB, andphlD, individually or in combination, identified a novel polyketide biosynthetic pathway in which PhlD is responsible for the production of monoacetylphloroglucinol (MAPG). PhlA, PhlC, and PhlB are necessary to convert MAPG to 2,4-DAPG, and they also may function in the synthesis of MAPG.

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